While recent advances in science and technology are helping to improve quality and add years to human life, the prevention of atherosclerosis, the underlying cause of cardiovascular disease (“CVD”) has not been sufficiently addressed. Atherosclerosis is a degenerative process resulting from an interplay of inherited (genetic) factors and environmental factors such as diet and lifestyle. Research to date suggest that cholesterol may play a role in atherosclerosis by forming atherosclerotic plaques in blood vessels, ultimately cutting off blood supply to the heart muscle or alternatively to the brain or limbs, depending on the location of the plaque in the arterial tree1,2. Data from the early Framingham Epidemiological Study indicates that increases in serum cholesterol levels are associated with increased risk of death from CVD3. More recent studies confirm that CVD is a leading cause of death and disability in industrialized nations4.
Studies have indicated that a 1% reduction in a person's total serum cholesterol yields a 2% reduction in risk of a coronary artery event5. Statistically, a 10% decrease in average serum cholesterol (e.g. from 6.0 mmol/L to 5.3 mmol/L) may result in the prevention of 100,000 deaths in the United States annually6.
As the population becomes increasingly aware of the importance of maintaining cholesterol balance in check, the need for naturally derived, safe and effective agents which address the underlying causes of CVD, and which can be readily incorporated into a wide variety of delivery means, becomes even more apparent.
One focus of such research related to naturally derived, safe and effective agents to address the underlying causes of CVD has been plant-derived sterols and stanols (also known as phytosterols and phytostanols). Sterols are naturally occurring compounds that perform many critical cellular functions. Phytosterols such as campesterol, stigmasterol and beta-sitosterol in plants, ergosterol in fungi and cholesterol in animals are each primary components of cellular and sub-cellular membranes in their respective cell types. The dietary source of phytosterols in humans comes from plant materials i.e. vegetables and plant oils. The estimated daily phytosterol content in the conventional western-type diet is approximately 60-80 milligrams in contrast to a vegetarian diet which would provide about 500 milligrams per day.
Phytosterols have received a great deal of attention due to their ability to decrease serum cholesterol levels when fed to a number of mammalian species, including humans. While the precise mechanism of action remains largely unknown, the relationship between cholesterol and phytosterols is apparently due in part to the similarities between the respective chemical structures (the differences occurring in the side chains of the molecules). It is assumed that phytosterols displace cholesterol from the micellar phase and thereby reduce its absorption or possibly compete with receptor and/or carrier sites in the cholesterol absorption process.
Over forty years ago, Eli Lilly marketed a sterol preparation from tall oil and later from soybean oil called Cytellin™ which was found to lower serum cholesterol by about 9% according to one report7. Various subsequent researchers have explored the effects of sitosterol preparations on plasma lipid and lipoprotein concentrations8 and the effects of sitosterol and campesterol from soybean and tall oil sources on serum cholesterols.9 Compositions have been explored in which phytosterols or phytostanols (their hydrogenated counterparts) are esterified in order to enhance solubility. One composition of phytosterols which has been found to be highly effective in lowering serum cholesterol is disclosed in U.S. Pat. No. 5,770,749 to Kutney et al.
Despite the obvious and now well recorded advantages of phytosterols, not only in the treatment of CVD and its underlying conditions such as hypercholesterolemia, hyperlipidemia, atherosclerosis, hypertension, thrombosis but in the treatment of other diseases such as Type II diabetes, dementia cancer and aging, the administration of phytosterols and the incorporation thereof into foods, pharmaceuticals and other delivery vehicles has been complicated by the fact that they are highly hydrophobic (i.e. they have poor water solubility). This highly hydrophobic nature of phytosterols renders them insoluble and barely dispersible in aqueous media. As such, phytosterols tend to be added to the fat phase of fat-based food products. Health-conscious consumers wishing to benefit from the cholesterol lowering effects of phytosterols are therefore forced to consume fat-rich foods, despite the health risks of a high fat diet.
In addition, and critically in the area of food and beverage production, phytosterols have a waxy consistency and a high melting point, creating solubility issues for the food processor. While they are oil-dispersible to some extent in their raw form, the amount required to produce an efficacious effect in a finished product can cause granulation. The current answer to this problem is esterification, which creates something of an equilibrium between the phytosterols and liquid oil. Due to these physical property limitations of phytosterols, their use by food manufacturers has been limited to fat-based products like margarine, salad dressings and, most recently, snack bars.
Furthermore, studies have investigated how the form (for example crystalline, suspension, granular) in which the phytosterols are dosed impacts on their ability to lower serum cholesterol levels. As phytosterols are highly hydrophobic, they do not dissolve to any appreciable extent in the micellar phase in the digestive tract and therefore are not capable of efficiently blocking cholesterol absorption. Oils and fats are capable to a limited but not satisfactory degree of dissolving free phytosterols. Only substantially solubilized phytosterols appear inhibit the absorption of cholesterol.
As noted above, since phytosterols have high melting points (typically about 136-150° C.) it is important to maintain a temperature of 80° C. or higher during dissolution of phytosterols in fats or oils, in order to avoid recrystallization of the phytosterols. Crystalline phytosterol imparts an unpleasant grainy, waxy texture to edible and topical products. However, at 80° C. and above commonly used fats and oils are vulnerable to oxidation. Rancid oils and fats detract greatly from the organoleptic properties of food products in particular. Hence there is a need to address the issue of this waxy texture in order to make any deliverable foods and beverages palatable and marketable.
The problems associated with adding phytosterols to the fat phase are compounded in low fat, fat-based products and in non-fat products. The amount of phytosterol capable of being dispersed in the fat phase of a fat-based emulsified product directly correlates with the amount of lipid in the product. Thus, when the lipid content is reduced to below a certain level it becomes technically impossible to incorporate enough phytosterol into an edible product to obtain a tangible health benefit. The problems posed by the need to disperse phytosterols in fat become more acute the lower the fat content of a fat-based product.
Early research on phytosterols focused on grinding or milling the phytosterols in order to enhance their solubility (U.S. Pat. Nos. 3,881,005 and 4,195,084 both to Eli Lilly). In addition, researchers have looked to the esterification of phytosterols in order to enhance their solubility. German Patent 2035069/Jan. 28, 1971 (analogous to U.S. Pat. No. 3,751,569) describes the addition of phytosterol fatty acid esters to cooking oil. The esterification is carried out between a free sterol and a fatty acid anhydride, with perchloric acid as the catalyst. The significant drawback to this process, along with others, is the use of non-food grade catalysts and reagents.
Conventionally, phytosterols have been incorporated into food products by melting a sterol or stanol, incorporating it into an oil phase, and blending the oil phase with other components to result in a phytosterol-containing food product. However, the aforementioned high melting points can result in significant crystallization of the phytosterols within the oil phase of such food products. Such crystallization results in food products with a gritty and unacceptable texture. This gritty texture is especially detectable when the oil/plant sterol phase is incorporated at high levels in the food product. The high melting points and hydrophobic nature of such phytosterols also make it difficult to blend such them with an aqueous phase. Furthermore, actual melting of the plant sterol for incorporation into food products is energy intensive.
Attempts have been made to solve these problems using, for example, chemical modification of the phytosterols. For example, as noted above, esterification of phytosterols generally results in lowered melting temperatures. Thus, such phytosterol esters generally may be incorporated into food products more readily due to the lower melting points and can provide food products without significantly gritty texture. Although the problem of fat solubility of phytosterols can be improved by esterification, this is not a completely satisfactory solution to the problem for two reasons: 1) phytosterol esters are biologically less effective than non-esterified phytosterols; and 2) when a phytosterol or phytosterol ester is distributed within the small volume lipid phase of a low fat emulsified product, the taste of the product is adversely affected, since the high concentration of phytosterol in the fat leads to a waxy sensation in the mouth and on the tongue.
Within the last 6-8 years, several different approaches have been used to incorporate plant sterols into food products. For example, European Patent Application EP 0 896 671 A (published Feb. 24, 1999) provides an aqueous dispersion of phytosterols by melting the phytosterols and emulsifiers to form a molten mixture and then dispersing the molten mixture in water using high shear. It was reported “that the step of melting the high melting phytosterols with surfactant prior to dispersion in water with or without surfactant contributes importantly to the ability to prepare a very fine dispersion with the use of high shear mixing or homogenization of the phytosterol or othermelting lipid”. The phytosterols are reported to have particle sizes of less than 15 microns and preferably less than 10 microns in aqueous dispersions. Such phytosterol dispersions could be incorporated into food products without the grittiness normally associated with phytosterols.
In WO2003105611, Coca Cola attempted to address the issue of incorporating highly hydrophobic compounds, such as phytosterols, into fruit beverages. It is noted in the application that “since hydrophobic ingredients have a different density than water and as a result, at the time of purchase and consumption of the product, a hydrophobic component may separate and float to the surface or sink to the bottom. The hydrophobic component that floats to the surface produces undesirable “ringing,” which is found in beverages, such as juices containing a hydrophobic ingredient with a density less than water, and results in a product that is non-uniform throughout the container.” Coca Cola has attempted to solve this problem by creating aqueous sterol dispersions without any added emulsifiers or thickening agents.
Earlier researchers attempted to overcome formulation limitations in relation to phytosterols by using several methods including: homogenization, encapsulation, and/or the addition of stabilizers, gums and the like. However, these methods increase the cost of the product, and in some instances are illegal in certain standardized products such as citrus juices.
Tiainen et al., U.S. Pat. No. 6,129,944 describes a method for producing a product containing a plant sterol by forming a homogeneous suspension of a microcrystalline plant sterol and a sweetening agent in an aqueous solution.
Vulfson et al., WO 00/41491 discloses hydrophobic compounds such as plant sterols and lycopenes as supplements to food products and beverages such as oleomargarine products, drinks, soups, sauces, dips, salad dressings, mayonnaise, confectionery products, breads, cakes, biscuits, breakfast cereals and yogurt type products. Vulson et al., in combining the plant sterol or lycopene with the food product, theorizes that the food product which has both hydroxyl and carboxyl groups interacts with the surface of the sterol or lycopene.
Haarasilta et al., WO 98/58554, describes a premix used in the food industry containing a pulverized plant sterol and a conventional foodstuff ingredient such as fruit, vegetable or berry type of material, particularly in a powder form and methods for preparing the premix.
Zawistowski, WO 00/45648, describes a method of preparing microparticles of plant sterols and plant stanols or mixtures of both by dispersing and suspending the plant sterols and plant stanols in a semi-fluid, fluid or viscous vehicle and exposing the vehicle so formed to impact forces. The method involves dispersing or otherwise suspending the plant sterol and/or plant stanol in a suitable semi-fluid, fluid or viscous vehicle followed by applying impact forces to the vehicle to produce microparticles. Zawistowski develops these impact forces by creating high-shear either with an air atomization nozzle, a pneumatic nozzle, a high-shear mixer, or colloid mill, but preferably a microfluidizer commercially available from Microfluidics Incorporation, Newton, Mass. Zawistowski observed that the plant sterols and/or plant stanols prepared in this way have greater “solubility” not only in oil based delivery systems but also in other media and can be incorporated into beverages such as colas, juices or dietary supplement and/or milk replacement drinks.
In order to increase the solubility of plant sterol, some researchers have synthesized various derivatives of plant sterol. For example, sitosterol mixed in certain ratios with starch hydrolysate, silicon dioxide and polyoxylene sorbitan monostearate through homogenizing, deaeration, pasteurizing and evaporation steps to form a medicinal powder for oral application, as disclosed in U.S. Pat. No. 3,881,005.
U.S. Pat. No. 5,932,562 discloses an aqueous homogeneous micellar mix of a plant sterol, lecithin and lysolecithin which has been dried to a finely divided water soluble powder. Other water-soluble plant sterols can be found in U.S. Pat. Nos. 6,054,144 and 6,110,502. According to these patents, aqueous-dispersible plant sterol is produced by admixing oryzanol or plant sterol, a monofunctional surfactant and polyfunctional surfactant in water at fixed ratios, and drying the admixture. This production method is characterized by being free from homogenization and deaeration steps with adoption of polyoxylene sorbitan monopalmitate and sorbitan monopalmitate as a monofunctional surfactant and a polyfunctional surfactant, respectively.
U.S. Pat. No. 6,190,720 discloses a food ingredient that can be used as a cholesterol-lowering agent, teaching that the food ingredient can be prepared by combining one or more molten plant sterols with one or more fats and one or more emulsifiers to homogeneity and cooling the homogeneous mixture to about 60° C. under agitation to give a paste. This food ingredient can be applied to oil-based foods such as salad dressings, margarine, etc.
PCT WO 00/33669 teaches that plant sterols can be dissolved or mixed in a melt of a food emulsifier, admixed with protein-containing foods such as milk or yogurt, homogenized, and added to food products. The dispersion stability of the cholesterol reducing, edible products is maintained only in the presence of a protein-containing material.
U.S. Pat. No. 6,267,963 describes a plant sterol-emulsifier complex which has a melting temperature at least 30° C. below that of the plant sterol, characterized in that, due to its reduced melting temperature, the plant sterol-emulsifer is less likely to crystallize during or after the manufacture of food products, and can be incorporated into food products in an amount effective to reduce serum cholesterol levels in a human consuming the food products without unpleasant effects on the texture of the food products.
In view of the technical difficulties involved in adding phytosterols to foods and beverages, and bearing in mind the utility of being able to widely supplement a wide variety of comestible products with these components, it would be highly advantageous to find an effective means of dispersing or suspending phytosterols in aqueous media at high concentrations or in creating a means to deliver phytosterols in a manner which addresses the problems of waxiness and guminess attendant in the powder formulation, thereby opening up the possibility of providing low fat or fat-free products (i.e. aqueous based products) containing phytosterols in a variety of formats.
It is an object of the present invention to obviate or mitigate the above noted disadvantages and to find a solution for the problem plaguing manufacturers wishing to widely use hydrophobic compounds, such as phytosterols, in these varied formats.